Abstract A04: Novel indol-based tambjamine analogs down-regulate survivin in lung cancer

Abstract

Abstract Introduction: Lung cancer is the most frequent cause of cancer death worldwide, with an increasing incidence and a poor prognosis that depends on tumor stage at presentation. Characterization of molecular abnormalities that play a crucial role on lung cancer will allow the development of targeted therapies for this illness. Our aim is to test novel indol-based compounds chemically related to tambjamines and to characterize the alterations in signaling pathways caused by their treatment in order to identify their molecular targets. Materials and Methods: Cell lines representative of the four major groups of lung cancer types were selected to perform this study: A549 (adenocarcinoma), SW900 (squamous cell carcinoma), H460 (large cell carcinoma) and DMS53 (small cell lung carcinoma). Cell viability evaluation was performed by the standard MTT assay. In order to characterize the main transcriptional changes produced by the selected compounds, a QIAGEN array of genes related to human cancer drug targets was used. Transcriptional alteration of a set of genes was further corroborated by TaqMan gene expression assays in order to validate the array. Additionally, the protein expression levels of several apoptotic-related proteins were assessed by immunoblot in the A549 cell line exposed to IC25, IC50 and IC75 values of the selected compounds. Moreover, Z-VAD-FMK (a pan-caspase inhibitor) treatment was performed and the outcome analyzed using the MUSE cell count and viability assay kit in order to assess the reversion in viability upon compounds treatment. Lastly, the p38 specific inhibitor SB202190 was used to assess its potential reversion of survivin levels and apoptosis by Western blot. Results and Discussion: Cell viability was significantly reduced in a dose-dependent manner by tambjamine analogs in all the lung cancer cell lines studied. Our results, using gene expression arrays, showed the impact of our compounds in several targets from apoptosis (e.g. BCL2 and BIRC5/survivin) and cell cycle (e.g. AURKs, CDKs and PLKs), among others. The down-regulation of the anti-apoptotic proteins survivin and Bcl2, as well as the activation of several pro-apoptotic proteins (initiator caspases, effector caspases and final executor PARP protein) was analyzed by immunoblot. Furthermore, the inhibition of caspase activity significantly reversed the cytotoxic effect of the selected compounds. Finally, the p38 stress kinase was activated after compound treatment and experiments using a specific inhibitor showed to partially recover the down-regulation of survivin levels and the activation of apoptosis. All these results hint that after compounds treatment cell death process is triggered mainly by apoptosis and that survivin down-regulation might be the responsible for the induction of this process. Conclusion: Altogether, our selected compounds show more potent cytotoxic properties than the first-line treatment cisplatinum, altering the expression of several key signaling proteins in lung cancer cells, especially apoptotic related proteins. These compounds strongly down-regulate the survivin cellular pool thereby triggering apoptotic cell death via caspase activation. Acknowledgements: This work was supported by the grant from the Spanish government and the EU (FIS PI13/00089) and a grant from La Marató de TV3 Foundation (20132730). LKG holds a FCT (SFRH/BPD/91766/2012) fellowship. Citation Format: Pilar Manuel-Manresa, Luis Korrodi-Gregório, Ananda Marina Rodilla, Roberto Quesada, Vanessa Soto-Cerrato, Ricardo Pérez-Tomás. Novel indol-based tambjamine analogs down-regulate survivin in lung cancer. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Targeting the Vulnerabilities of Cancer; May 16-19, 2016; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(1_Suppl):Abstract nr A04.